Cockroaches (CR) infesting houses produce protein allergens which are an important cause of IgE mediated allergic reactions and asthma among minority populations living in sub-standard housing. The aims of this grant are to clone allergens from Blattella germanica and Periplaneta americana; to produce recombinant B. germanica allergens (Bla g 2, Bla g 4 and Bla g 5); to develop immunoassays to assess exposure to CR; to produce allergen molecules that no longer bind IgE; and to analyze T cell responses to CR allergens. These aims will be achieved by screening cDNA libraries with IgE antibodies (ab) and producing recombinant allergens in expression vectors. The biologic activity of recombinant Bla g 2, Bla g 4 & Bla g 5 will be compared by skin testing, histamine release and serum IgE ab assays in a multi-center study of patients with asthma enrolled in Charlottesville, New Orleans, Baltimore, Augusta, GA, and in Strasbourg, France. Exposure to CR allergens in houses will be assessed using monoclonal antibody based assays and counts of CR infestation. These experiments will focus on developing tests for P. americana allergens, as well as B. germanica, in order to define allergens levels in dust samples which cause IgE sensitization and symptoms. Also, to investigate the aerodynamic properties of CR allergens. Site directed mutagenesis will be used to localize IgE ab sites on Bla g 4 and Bla g 5 and to generate structural alterations that reduce binding to IgE. Amino acids will be targeted for mutagenesis by molecular modeling. T cell responses to recombinant allergens will be compared by proliferation assays and the prevalence and magnitude of responses to each allergen will be determined. T cell sites will be mapped using allergen variants, synthetic peptides or expressed partial allergen sequences. This grant will increase our understanding of the role and importance of specific CR allergens in causing asthma. The production of altered allergens by mutagenesis, combined with analyses of T cell responses, may also provide more rational strategies for immunotherapy of CR asthma.